Bordetella pertussis (B. pertussis) is the etiological agent of pertussis, a disease of the respiratory tract which is transmitted from a sick person to a susceptible healthy individual during the virulent stage.
Pertussis, which is characterised by convulsive coughing fits and serious respiratory symptomology, strikes individuals of all ages and, in the first years of life, may cause death in 0.5% of cases.
The incidence of the infection can be controlled effectively by immunisation with a suitable vaccine.
Currently, a cellular anti-pertussis vaccine is used, that is, a vaccine constituted by entire virulent B.pertussis cells treated with merthiolate and killed at 56.degree. C. Whilst it confers protective immunity, this vaccine may, however, induces undesirable side effects ranging from simple wheals, erythema and fever to convulsions and brain damage. For these reasons, the use of this vaccine has been drastically reduced in recent years resulting in a new explosion of pertussis cases.
As a result, there is a need to develop an anti-pertussis vaccine which does not have the disadvantages described above.
It is known that, during the virulent stage (Stage I), B.pertussis produces a series of toxic components (virulence factors) which are necessary in order for the infection to arise and persist, and some of which seem to be immunogens particularly suitable for the development of an acellular anti-pertussis vaccine.
Thus, for example, the pili or fimbriae, which are proteins present on the surface of the bacterium, and are constituted by polymerised (pilinic) subunits with molecular weights of from 21,000 to 24,000 Daltons, are involved in the specific adhesion of the bacteria to the cilia of the epithelial cells of the upper respiratory tract.
This is an essential stage in the pathogenisis of pertussis since it enables the microorganism to elude the host's defensive system. A vaccine against this stage would therefore be desirable.
Ashworth et al (1982) (Infect. Immun., 37, 1278-1281) first suggested that the pili of B.pertussis were sertotype-specific agglutinogens, that is surface antigens which stimulate the production of antibodies which agglutinate bacterial cells of the same serotype.
Currently, antigenically different pilinic proteins belonging to serotypes 1, 2, 3, 4, 5, 6 and x have been identified.
The use of purified pili for the preparation of a vaccine must, however, take account of the antigenic variation observed in strains of B.pertussis.
In fact it has been found that, whilst some of these antigens are present in all the strains of B.pertussis, those associated with the fimbriae 2, 3, 4, 5 and 6 are present in different combinations not only in different strains, but also in different isolated samples of the same strain.
An ideal vaccine, that is a vaccine which can confer protective immunity against any infective strain of B.pertussis, should therefore contain all the serotypically different antigens.
The preparation of such a vaccine presents many problems resulting, on the one hand, from the fact that it is difficult to isolate and purify these antigens and, on the other hand, from their limited availability.
For these reasons, it would seem desirable to prepare a strain of Bordetella which can produce all the pilinic subunits with high yields.
The structural genes which code for the subunits fim2 and fimx of B.pertussis have recently been cloned and sequenced (Livey, J. et al., (1987), Mol. Microbiol., 1 (2), 203-209; Pedroni, P. et al., (1988), Mol. Microbiol., 2, 539-543).
Up to now, however, nothing has been known about the regulation of the expression of these genes and, in particular, the nucleotide sequences of their promoters were not known.